Effects of different ammonium sulfate concentrations on meilingmycin biosynthesis were studied in this research. The results show that a lower concentration of ammonium ions stimulates the biosynthesis of meilingmycin, while a concentration higher than 5mmol/L inhibits the mycelial growth and the biosynthesis of the products. However, increased sugar consumption rate with the elevated concentration of ammonium sulfate was observed during the fermentation process. On this basis, six enzymes, which are responsible for the meilingmycin biosynthesis and the glucose metabolism, were measured and analyzed during the bioprocess. The results suggest that glucose-6-phosphate dehydrogenase, citrate synthase, succinate dehydrogenase and fatty acid synthase are stimulated by higher concentration of ammonium ions, while valine dehydrogenase and methylmalonyl-CoA carboxyltransferase are inhibited. From the results it follows that the precursor supply was restricted with the higher concentration of ammonium ions, which results in the lower production of meilingmycin. Thus, the strategy of maintaining a low level of FAS activity is a critical factor for high yield of meilingmycin.